Cellular thiols are essential biomolecules that play important roles in biological systems as key components of protein structures and metabolic intermediates. The thiol molecules are critical in maintaining appropriate oxidation-reduction states of the proteins, cells and organisms. Glutathione (GSH), for example, is a thiol with the highest abundance in the cellular system. It is a small protein with a short sequence of Gly-Cys-Glu or GCE that resides in every cell of our body and is thus vital to life. Its deficiency results in cellular dysfunction. Many health problems, such as Alzheimer's disease, leucocyte loss, liver damage, psoriasis, cancer and AIDS are known to be associated with levels of cellular thiols.
The development of fluorescent probes for biomolecular detection has emerged as an active area of research due to its importance to bioscience and applications in biotechnology as well as its impact on public health. The fluorescent assay process offers a number of advantages over other analytical techniques, such as rapid response, high sensitivity, low background noise, and wide dynamic working range. Thanks to the enthusiastic effort of scientists devoted to this area of research, a large variety of fluorescent bioprobes have been developed. However, many of the bioprobes work in a “turn off” mode. For example, the emission of a fluorophore is switched “off” when it interacts with a quenching species in a biological system through a mechanism of fluorescence resonance energy transfer.
A sensitive and selective assay of thiols is thus of great biological implications. Much work has been done in the area of developing fluorescent sensors for thiol detection. In almost all the current sensing systems, thiols have been assayed by measuring the changes in the fluorescence signals of the probes with the analyte concentrations in the solution state using a spectrofluorometer. For real-world applications, however, it is preferable to perform the bioassays on solid strips because it requires no complex and expensive equipment and is thus simple, quick and convenient. No report has been found in the art about a thiol assay in the solid state to date.
A thorny problem often encountered by traditional fluorophores is aggregation-caused quenching (ACQ) of their emissions in the solid state. When dispersed in aqueous media or bound to biomolecules, the fluorophore molecules are inclined to aggregate, which usually quenches their fluorescence processes and thus greatly limits their effective ranges as bioprobes. The ACQ effect also makes it difficult to assay low-abundance molecular species in biological systems, because the fluorescence signals from the miniature amounts of the fluorophores matching the bioanalyte levels may be too weak to be determined accurately, while at the high fluorophore concentrations, the emissions are further weakened, rather than enhanced, due to the notorious ACQ effect.
An extraordinary phenomenon of aggregation-induced emission (AIE) that is exactly opposite to the ACQ effect discussed above, has been recently discovered. A series of fluorogen molecules that are non-emissive in solutions, such as tetraphenylethylene (TPE), are induced to emit efficiently by aggregate formation. The unique AIE effect has been utilized to develop new bioprobes of “turn on” type, which enjoy much higher sensitivity over their “turn off” counterparts. The selectivity of the AIE probes, however, is low and needs to be improved.
Accordingly, a new bioprobe with high selectivity while keeping its superb sensitivity has been desired in the art.